A method of analysis based on the complementarity of nucleic acid nucleotide sequence can be utilized to directly analyze genetic traits. Accordingly, this method of analysis is a very powerful means for identifying genetic diseases, canceration, microorganisms, etc. Further, because the gene itself is the object of detection, time-consuming and cumbersome procedures such as culture may be omitted.
However, since the detection of a target gene, which is present in a very small amount in a sample, is generally not easy, amplification of the target gene itself, its detection signal, or the like is required.
In the amplification of nucleic acids, the most general method for detecting the occurrence of amplification is carried out by subjecting the solution after amplification to agarose gel electrophoresis and binding a fluorescent intercalator such as ethidium bromide to the amplification product, thereby observing specific fluorescence. When there is no possibility of contamination by other DNA and only the occurrence of the amplification product is of interest, fluorescence can be observed by adding the fluorescent intercalator to the solution after amplification while omitting electrophoresis. While these methods are simple, a UV lamp and a darkroom are required to observe fluorescence.
Also, when amplification is performed using a primer or a nucleotide labeled with various label substances including a fluorescent dye, there is a method for detecting the label incorporated into the amplification product. This method, however, requires the separation of a free labeled primer (or nucleotide) that was not incorporated into the amplification product. Accordingly, this method is not suitable for the amplification of genes which uses very small amounts of reaction solution. Also, a labeled primer and nucleotide are expensive.